Type 1 Calreticulin Mutations Differentially Activate the IRE1α-XBP1 Pathway of the Unfolded Protein Response to Drive Myeloproliferative Neoplasms

Abstract Approximately 20% of patients with myeloproliferative neoplasms (MPN) harbor mutations in the gene calreticulin (CALR). 80% of CALR mutations are classified as either type 1 or type 2, exemplified by a 52 bp deletion (CALRdel52) and a 5 bp insertion (CALRins5), respectively. Despite their shared mutant C-termini and mutual ability to bind and activate MPL, patients with type 1 and type 2 CALR mutations display significant clinical and prognostic differences. Type 1 mutations are primarily associated with an MF phenotype and a higher risk of fibrotic transformation from ET, while type 2 mutations are more common in ET. Molecularly, type 2 CALR mutant proteins retain many of the calcium binding sites present in the wild type protein, while type 1 CALR mutant proteins lose these residues. The functional consequences of this differential loss of calcium binding sites remain yet unexplored. Current targeted therapies for CALR mutated MPN are not curative, and treatment does not differentiate between type 1 versus type 2 mutant CALR-driven disease, despite the different phenotypic and prognostic outcomes in these patients. In order to improve treatment strategies for CALR mutated MPN patients, it is critical to identify specific dependencies unique to each CALR mutation type that can be exploited for therapeutic gain. Here, we show that type 1 CALRdel52 but not type 2 CALRins5 mutations lead to activation of and dependency on the IRE1α-XBP1 pathway of the unfolded protein response (UPR). Mechanistically, we found that the loss of calcium binding residues in the type 1 mutant CALR protein directly impairs its calcium binding ability, which in turn leads to depleted ER calcium and subsequent activation of the IRE1α-XBP1 pathway. Using cell lines and primary MPN patient samples, we identified two novel transcriptional targets of XBP1 specific to type 1 CALRdel52-expressing cells - the anti-apoptotic protein BCL-2 and the calcium efflux channel IP3R. We show that BCL-2 acts downstream of XBP1 to promote survival in the face of depleted ER calcium, while IP3R is up-regulated downstream of XBP1 to promote continued ER calcium efflux in order to sustain IRE1α-XBP1 pathway activation and survival. We found that genetic or pharmacological inhibition of IRE1α-XBP1 signaling induced cell death only in type 1 mutant but not type 2 mutant or wild type CALR-expressing cells. Moreover, we show that in vivo inhibition of IRE1α significantly abrogates type 1 mutant CALR-driven disease in a bone marrow transplantation model, but has no effect on type 2 mutant CALR-driven disease. This work is the first to demonstrate that type 1 and type 2 mutant CALR-expressing cells display differential molecular dependencies that can be exploited for therapeutic gain. Moreover, this study answers an enduring question regarding the functional consequence of the loss of calcium binding sites on the type 1 mutant CALR protein, and demonstrates how type 1 CALR mutant-expressing cells rewire the UPR, downstream calcium signaling, and apoptotic pathways to drive MPN. Figure 1 Figure 1. Disclosures Koschmieder: BMS: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: (e.g. travel support); Shire: Honoraria, Other; Karthos: Other: Travel support; Ariad: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: (e.g. travel support); Incyte: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: (e.g. travel support); Geron: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: (e.g. travel support), Research Funding; Abbvie: Other: Travel support; Pfizer: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: (e.g. travel support); Alexion: Other: Travel support; Sanofi: Membership on an entity's Board of Directors or advisory committees, Other: Travel support; Baxalta: Membership on an entity's Board of Directors or advisory committees, Other; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: (e.g. travel support); CTI: Membership on an entity's Board of Directors or advisory committees, Other; Roche: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: (e.g. travel support), Research Funding; AOP Pharma: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: (e.g. travel support), Research Funding; Bristol-Myers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel support, Research Funding; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel support, Research Funding; Image Biosciences: Other: Travel support.

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Bleeding in Patients with Clinically Severe Von Willebrand Disease: Preliminary Results of Athn 9, a Natural History Study of the Safety, Effectiveness, and Practice of Treatment for People with Severe Von Willebrand Disease (VWD)

Blood ◽

10.1182/blood-2020-137124 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 27-28

Author(s):

Angela C. Weyand ◽

Kenneth D Friedman ◽

Sweta Gupta ◽

Kristina M. Haley ◽

Chunla He ◽

...

Keyword(s):

Board Of Directors ◽

Research Funding ◽

Von Willebrand Disease ◽

Heavy Menstrual Bleeding ◽

Severe Bleeding ◽

Advisory Committees ◽

Type 3 ◽

Von Willebrand

Background: Von Willebrand Disease (VWD) is the most common inherited bleeding disorder, thought to occur in ~0.1% of the population. VWD results from a quantitative (Type 1 or 3) or qualitative (Type 2) defect in von Willebrand Factor (VWF), a multifunctional plasma protein involved in primary and secondary hemostasis. Diagnosis of VWD can be difficult due to pre-analytical variables, a wide coefficient of variation in testing, and incomplete penetrance. Treatment of VWD is aimed at replacement of the defective or missing protein through plasma derived or recombinant VWF, release of endogenous VWF through desmopressin (DDAVP) or clot stabilization with anti-fibrinolytic therapy. Though individuals with mild VWD and bleeding symptoms are common, less is known regarding individuals with VWD and a clinically severe bleeding phenotype. Aims: To characterize the bleeding phenotype and treatment regimens in patients with clinically severe VWD in the United States. Study Design and Methods: ATHN 9 is sponsored by the American Thrombosis and Hemostasis Network (ATHN) and is being conducted at ATHN-affiliated sites across the US. Participants were identified by the site investigators with the projected goal to enroll 130 individuals. Inclusion criteria were patients with severe VWD defined as type 3 VWD, or VWF:RCo, VWF:GPIbM or VWF:Ag≤ 30% or patients with "clinically severe VWD" defined by VWF:RCo, VWF:GPIbM or VWF:Ag ≤ 40% a with severe bleeding phenotype (need for recurrent use of factor concentrates) and prior enrollment in the ATHN dataset national surveillance data collection project. Patients with platelet-type or acquired VWD were excluded. Laboratory assessment including a standardized diagnostic battery, VWF genetic analysis, and inhibitor testing, was performed by a central laboratory. Bleeding was assessed using the International Society for Thrombosis and Haemostasis (ISTH) Bleeding Assessment Tool (BAT) (normal adult 0-4, normal score <18 years 0-2) and the Pictorial Bleeding Assessment Chart (PBAC) if applicable. Results: Initial data on 35 participants was analyzed. Most were adult (69%), female (66%), Caucasian (86%) and non-Hispanic (89%). Just less than half (16/35, 45.7%) have completed diagnostic testing (Table 1). Half of the patients had Type 1, a quarter Type 3, and the remaining had Type 2 or unknown. The majority of patients (69%) had VWF GPIbM activity <30IU/dL, while 44% had an abnormally low FVIII level as well. The majority (26/35, 74.3%) had a known family history of VWD. Slightly over half (19/35, 54.2%) had previous surgery. Few participants (4/35, 11.4%) reported the presence of a target joint at enrollment, ankle being most common. The bleeding phenotype was significant but variable with a mean ISTH BAT score of 10.6 (range 0-39). With the exception of the youngest cohort (0-5 years of age, mean BAT score of 6, range 3-8), bleeding scores increased with age and all participants had abnormal scores. The most commonly endorsed symptoms were epistaxis, heavy menstrual bleeding (HMB), and post-surgical bleeding. The PBAC was performed on 4/10 participants in reference to their last period with a median score of 36 and range of 0-112 (>150 is abnormal). The majority (3/4) of participants filling out the PBAC received VWF concentrate prophylaxis for HMB. The majority (23 participants, 66%) utilized factor concentrates for prophylaxis or on-demand treatment; six patients (17%) were on continuous prophylaxis, while 12 (34%) were on event-based or HMB prophylaxis while the remainder received episodic treatment. Participants most commonly used plasma derived VWF concentrate (93.9%) with the remainder using recombinant VWF. Discussion: Initial evaluation of 35 participants with clinically severe VWD demonstrated a predominance of mucosal bleeding with a minority of participants endorsing joint bleeds at enrollment. Despite abnormal ISTH BAT scores in all participants, PBAC scores were within normal range, likely reflecting appropriate management of HMB with most participants receiving VWF concentrate for HMB prophylaxis. In contrast to patients with mild disease where antifibrinolytics and desmopressin are frequently used, factor replacement was the most common treatment modality. Future analysis will focus on laboratory evaluation, bleeding phenotype, response to factor replacement therapy and quality of life. Disclosures Weyand: Shire: Membership on an entity's Board of Directors or advisory committees; Kedrion: Membership on an entity's Board of Directors or advisory committees; Aptevo: Membership on an entity's Board of Directors or advisory committees; Bayer: Membership on an entity's Board of Directors or advisory committees. Friedman:Alexion: Speakers Bureau; Bayer: Consultancy; Instrumentation Laboratories: Consultancy; Alexion: Consultancy. Haley:ATHN: Research Funding. Roberts:Sanofi: Consultancy, Speakers Bureau; Novo Nordisk: Consultancy, Speakers Bureau; Pfizer: Consultancy; Takeda: Consultancy, Research Funding, Speakers Bureau; uniQure: Consultancy; Octapharma: Consultancy, Speakers Bureau. Sidonio:Genentech: Membership on an entity's Board of Directors or advisory committees; Sanofi/Bioverativ: Membership on an entity's Board of Directors or advisory committees; Octapharma: Membership on an entity's Board of Directors or advisory committees; Bayer: Membership on an entity's Board of Directors or advisory committees; Novo Nordisk: Membership on an entity's Board of Directors or advisory committees; Uniqure: Membership on an entity's Board of Directors or advisory committees; Biomarin: Membership on an entity's Board of Directors or advisory committees; Catalyst Sciences: Membership on an entity's Board of Directors or advisory committees; Emergent Solutions: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees; Pfizer: Membership on an entity's Board of Directors or advisory committees; Kedrion: Membership on an entity's Board of Directors or advisory committees; Takeda: Research Funding; Genentech: Membership on an entity's Board of Directors or advisory committees; Octapharma: Research Funding; Grifols: Research Funding; Grifols: Membership on an entity's Board of Directors or advisory committees. OffLabel Disclosure: VWF concentrates used for heavy menstrual bleeding

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Differential Impact of Interferon Alpha on JAK2V617F and Calr Mutated Hematopoietic Stem and Progenitor Cells in Classical MPN

Blood ◽

10.1182/blood-2018-99-114744 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 4333-4333 ◽

Cited By ~ 1

Author(s):

Matthieu Mosca ◽

Lamia Lamrani ◽

Christophe Marzac ◽

Amandine Tisserand ◽

Valérie Edmond ◽

...

Keyword(s):

Board Of Directors ◽

Interferon Alpha ◽

Research Funding ◽

Molecular Response ◽

Advisory Committees ◽

Hematopoietic Stem ◽

Clonal Architecture

Abstract Introduction: Classical BCR-ABL-negative myeloproliferative neoplasms (MPN) include Polycythemia Vera (PV), Essential Thrombocytemia (ET) and Primary Myelofibrosis (PMF). They are acquired clonal disorders of hematopoietic stem cells (HSC) leading to the hyperplasia of one or several myeloid lineages. They are due to three main recurrent mutations affecting the JAK/STAT signaling pathway: JAK2V617F and mutations in the calreticulin (CALR) and thrombopoietin receptor (MPL). Interferon alpha (IFNα) is the only drug that not only induces a hematological response in ET, PV and early MF, but also a significant molecular response on both JAK2V617F or CALR-mutated cells. Our broad aim was to understand the mechanism of action of IFNα. Previously, our group and others have shown that IFNα specifically targets JAK2V617F HSC in a chimeric JAK2V617F knock-in mouse model. In this study, we wanted to know how and how fast IFNα impacts the different mutated human hematopoietic compartments. Methods: A prospective study was performed with a cohort of 47 patients treated by IFNα for 3-5 years. The MPN disease distribution was 40% ET, 49% PV and 11% MF. This cohort included 33 JAK2V617F-mutated patients, 11 CALR-mutated patients (7 type 1/type 1-like and 4 type 2/type 2-like), 2 both JAK2V617F- and CALR-mutated patients and 1 MPLW515K-mutated patient. At 4-month intervals, the JAK2V617F or/and CALR mutation allele frequency was measured in mature cells (granulocytes, platelets). Simultaneously, the clonal architecture was also determined by studying the presence of the JAK2V617F or CALR mutations in colonies derived from the different hematopoietic stem and progenitor cell (HSPC) populations (CD90+CD34+CD38- HSC-enriched progenitors, CD90-CD34+CD38- immature progenitors and CD90- CD34+CD38+ committed progenitors). Results: After a median follow-up of 33 months, IFNα targets more efficiently and rapidly the HSPC particularly in HSC-enriched progenitors, than the mature blood cells in JAK2V617F patients (p<.05). Moreover, homozygous JAK2V617F clones responded more rapidly than heterozygous clones in all hematopoietic cell compartments showing that the intensity of JAK2V617F signaling is correlated with the efficacy of IFNα. This efficacy was slightly increased after a median follow-up of 51 months. In contrast, during a median follow-up of 33 months for CALR-mutated patients, IFNα targeted similarly the HSPC and the mature cells. Moreover, IFNα induced a slower response in targeting CALR-mutated HSPC than the JAK2V617F HSPC (p<.05) (see Figure). The role of associated mutations at diagnosis was also investigated in the IFNα-mediated HSPC molecular responses using a NGS targeted myeloid panel. In JAK2V617F-mutated patients, the number of associated mutations did not impact the HSPC molecular response. In contrast, in CALR-mutated patients, the only molecular responders were not associated with other mutations, although the lower number of cases should be expanded. Using Ba/F3-MPL cellular models and primary cells, we observed that JAK2V617F was more prone to sensitize to IFNα signaling (increased Phospho-STAT1 and IFN-stimulating genes (ISGs)) compared to controls or CALRdel52 mutated cells. Conclusion: Altogether, our results show that IFNα targets more efficiently the human JAK2V617F-HSPCthan the mature cells. Moreover, IFNα has a greater efficacy on JAK2V617F HSPC thanCALR-mutated HSPC. This former result was associated with a greater priming of the IFNα signaling by JAK2V617F than by CALRdel52. The molecular response was dependent not only on mutational status, but also on the presence of other associated mutations for the CALR-mutated HSPC. Patient data are currently incorporated into a mathematical model taking into account clonal architecture and associated mutations to develop an algorythm able to predict patient response. Figure. Figure. Disclosures Kiladjian: Celgene: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; AOP Orphan: Membership on an entity's Board of Directors or advisory committees, Research Funding.

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Profiling T-Cell Receptor Diversity and Dynamics during Lymphoma Immunotherapy Using Cell-Free DNA (cfDNA)

Blood ◽

10.1182/blood-2020-141655 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 49-50

Author(s):

Navika D Shukla ◽

Alexander F. M. Craig ◽

Brian Sworder ◽

David M. Kurtz ◽

Charles Macaulay ◽

...

Keyword(s):

T Cell ◽

Cell Therapy ◽

Board Of Directors ◽

Research Funding ◽

Advisory Committees ◽

T Cell Therapy ◽

Car T Cell ◽

Car T ◽

Tcr Repertoire ◽

Support Research

Background: Characterization of T-cell receptor (TCR) diversity and dynamics is increasingly critical to understanding therapeutic immune responses targeting tumors. Current TCR profiling methods generally require invasive tissue biopsies that capture a single snapshot of immune activity or are limited by the sheer diversity of the circulating TCR repertoire. In theory, T-cells with the greatest turnover could best reflect pivotal immune dynamics from both circulating and tissue-derived compartments, including non-circulating tissue-resident memory T-cells (Trm). To noninvasively capture such responses in the blood, we developed and benchmarked a high-throughput TCR profiling approach using plasma, optimized for the fragmented nature of cfDNA and the non-templated nature of rearranged TCRs. We then applied this method for residual disease monitoring in mature T-cell lymphomas (TCL) without circulating disease and for characterizing immune dynamics after anti-CD19 chimeric antigen receptor (CAR19) T-cell therapy of B-cell lymphomas with axicabtagene ciloleucel. Methods: We developed SABER (Sequence Affinity capture & analysis By Enumeration of cell-free Receptors) as a technique for TCR enrichment and analysis of fragmented rearrangements shed in cfDNA and applied this method using Cancer Personalized Profiling by Deep Sequencing (CAPP-Seq). We used SABER to profile a total of 381 samples (300 cfDNA and 81 PBMC samples) from 75 lymphoma patients and 18 healthy controls. After mapping sequencing reads (hg38) to identify candidate rearrangements within TCR loci, unique cfDNA fragments were resolved by a novel strategy to define consensus of unique molecular identifiers clustered by Levenshtein distances, followed by CDR3-anchoring for enumeration of final receptor clonotypes. SABER thus leverages information from fragmented TCRs, a critical requirement for cfDNA, to make V gene, CDR3, and J gene assignments after deduplication-mediated error-correction. We benchmarked SABER against established amplicon-based TCR-β targeted sequencing (LymphoTrack, Invivoscribe) and repertoire analysis methods (MiXCR; Bolotin et al, 2015 Nature Methods) when considering both cfDNA and PBMC samples from healthy adults and TCL patients. We assessed SABER performance for tracking clonal molecular disease in patients with mature TCLs from both cellular and cell-free circulating compartments (n=9). Malignant TCL clonotypes were identified in tumor specimens using clonoSEQ (Adaptive Biotechnologies). Finally, we evaluated TCR repertoire dynamics over time in 66 DLBCL patients after CAR19 T-cell therapy. Results: SABER demonstrated superior recovery of TCR clonotypes from cfDNA compared to both amplicon sequencing (LymphoTrack, Invivoscribe) and hybrid-capture methods when enumerating receptors using MiXCR (Fig. 1A). When applied to blood samples from TCL patients, SABER identified the malignant clonal TCR-β rearrangement in 8/9 (88.9%) cases, with significantly improved detection in cfDNA (p=0.015, Fig. 1B). Specifically, tumoral TCR clonotype was detectable only in cfDNA in 6 cases (75%), cfDNA-enriched in 1 case (12.5%), and detectable only in PBMCs in 1 case (12.5%). We applied SABER to monitor TCR repertoire dynamics in cfDNA after CAR T-cell therapy of patients with relapsed/refractory DLBCL and observed increased T-cell turnover and repertoire expansion (greater total TCR-β clonotypes) (Fig. 1C). As early as 1-week after CAR19 infusion, TCR repertoire size was significantly correlated both with cellular CAR19 T-cell levels by flow cytometry (p=0.008) as well as with retroviral CAR19 levels in cfDNA (p=2.20e-07) suggesting faithful monitoring of CAR T-cell activity (Fig. 1D). TCR repertoire size one month after infusion was significantly associated with longer progression-free survival (HR 0.246, 95% CI 0.080-0.754, p=0.014). Conclusions: SABER has a favorable profile for cfDNA TCR repertoire capture when compared to existing methods and could thus have potential broad applicability to diverse disease contexts. Given the higher abundance of lymphoma-derived TCRs in cfDNA than intact circulating leukocytes, SABER holds promise for monitoring minimal residual disease in T-cell lymphomas. This approach also holds promise for monitoring T-cell repertoire changes including after CAR T-cell therapy and for predicting therapeutic responses. Disclosures Kurtz: Genentech: Consultancy; Foresight Diagnostics: Other: Ownership; Roche: Consultancy. Kim:Corvus: Research Funding; Eisai: Membership on an entity's Board of Directors or advisory committees, Research Funding; Elorac: Research Funding; Forty Seven Inc: Research Funding; Galderma: Membership on an entity's Board of Directors or advisory committees, Research Funding; Horizon Pharma: Consultancy, Research Funding; Innate Pharma: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Kyowa-Kirin Pharma: Research Funding; Medivir: Membership on an entity's Board of Directors or advisory committees; Merck: Research Funding; miRagen: Research Funding; Neumedicine: Consultancy, Research Funding; Portola: Research Funding; Seattle Genetics: Membership on an entity's Board of Directors or advisory committees; Solingenix: Research Funding; Takeda: Membership on an entity's Board of Directors or advisory committees, Research Funding; Trillium: Research Funding. Mackall:Lyell Immunopharma: Consultancy, Current equity holder in private company; BMS: Consultancy; Allogene: Current equity holder in publicly-traded company; Apricity Health: Consultancy, Current equity holder in private company; Nektar Therapeutics: Consultancy; NeoImmune Tech: Consultancy. Miklos:Kite-Gilead: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Travel support, Research Funding; Adaptive Biotech: Consultancy, Other: Travel support, Research Funding; Juno-Celgene-Bristol-Myers Squibb: Consultancy, Other: Travel support, Research Funding; Novartis: Consultancy, Other: Travel support, Research Funding; Allogene Therapeutics Inc.: Research Funding; Pharmacyclics: Consultancy, Other: Travel support, Patents & Royalties, Research Funding; Janssen: Consultancy, Other: Travel support; Miltenyi Biotec: Research Funding. Diehn:Varian Medical Systems: Research Funding; Illumina: Research Funding; Roche: Consultancy; AstraZeneca: Consultancy; RefleXion: Consultancy; BioNTech: Consultancy. Khodadoust:Seattle Genetics: Consultancy; Kyowa Kirin: Consultancy. Alizadeh:Janssen: Consultancy; Genentech: Consultancy; Pharmacyclics: Consultancy; Chugai: Consultancy; Celgene: Consultancy; Gilead: Consultancy; Roche: Consultancy; Pfizer: Research Funding.

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Promoter-Wide Transcriptional Deregulation In Waldenstrom Macroglobulinemia

Blood ◽

10.1182/blood.v116.21.3620.3620 ◽

2010 ◽

Vol 116 (21) ◽

pp. 3620-3620

Author(s):

Yang Liu ◽

Min Ni ◽

Aldo M. Roccaro ◽

Xavier Leleu ◽

Yong Zhang ◽

...

Keyword(s):

Gene Expression ◽

Bone Marrow ◽

Board Of Directors ◽

Binding Sites ◽

Research Funding ◽

Enrichment Analysis ◽

Promoter Regions ◽

Advisory Committees ◽

Pol Ii ◽

Normal Controls

Abstract Abstract 3620 Introduction: Waldenstrom macroglobulinemia (WM) is a rare indolent non-Hodgkin lymphoma, characterized by bone marrow infiltration of clonal lymphoplasmacytic cells. Despite recent advances in understanding the pathogenesis of this disease, the molecular basis of WM etiology has not been clearly defined. We therefore performed genome-wide analysis of RNA polymerase II (pol II) binding sites and gene expression profiling in primary WM cells in order to comprehensively define the aberrant transcriptional regulation and related genes in WM. Methods: Primary CD19+ bone marrow derived WM cells and normal primary bone marrow were used. Genomic DNA was extracted using genome isolation kit (QIAGEN) after cross linking. All the DNA samples were sent for Chip assay and human promoter 1.0R array (Genepathway Inc.) which comprised of over 4.6 million probes tiled through over 25.500 human promoter regions. Each promoter region covers approximately 7.6kb upstream through 2.45kb downstream of the transcription start sites. For over 1,300 cancer associated genes, coverage of promoter regions was expanded to additional genomic content; for selected genes total coverage spans from 10kb upstream through 2.45kb downstream of transcription start sites. The published gene expression datasets (GDS2643) which included 10 CD19+ B cell from bone marrow of 10 WM patients and 8 normal controls was analyzed by d-chip software and normalized to normal control. The motif analysis was performed using Cistrome online tools from the Dana Farber Cancer Institute. The gene sets enrichment analysis (GSEA) was performed using GSEA online software from Broad institute. Results: A total of 13,546 high-confidence pol II sites were identified in WM samples and share a small percentage of overlap (11.5%) with the binding sites identified in normal controls. Combining the expression microarray data of WM patient samples and normal controls, we demonstrated a significant correlation between high levels of gene expression and enriched promoter binding of pol II. Notably, we also observed that the WM-unique pol II binding sites are localized in the promoters of 5,556 genes which are involved in important signaling pathways, such as Jak/STAT and MAPK pathways by applying gene set enrichment analysis (GSEA). Interestingly, we found that STAT, FOXO and IRF family binding sites motifs were enriched in the pol II-bound promoter region of IL-6 which plays a crucial role in cell proliferation and survival of WM cells. Moreover, the CpG island associated c-fos promoter was enriched for Pol II binding as compared to the normal control. Conclusion: The presence of increased Pol II binding and the identification of transcription factor motifs in the promoters of key oncogenes may lead to a better understanding of WM. Our findings suggest that altered transcriptional regulation may play an important role in the pathogenesis of WM. In addition, this study will provide novel insights into the molecular mechanism of WM etiology, and may lead to discovery of novel diagnostic molecular biomarkers and therapeutic targets for WM. Disclosures: Leleu: Celgene: Consultancy, Research Funding; Janssen Cilag: Consultancy, Research Funding; Leo Pharma: Consultancy; Amgen: Consultancy; Chugai: Research Funding; Roche: Consultancy, Research Funding; Novartis: Consultancy, Research Funding. Ghobrial:Celgene: Membership on an entity's Board of Directors or advisory committees; Millennium: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees.

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Improvements in Skeletal Manifestations in Gaucher Disease Type 1 Patients After 3 Years of Treatment with Oral Eliglustat During a Phase 2 Trial,

Blood ◽

10.1182/blood.v118.21.3216.3216 ◽

2011 ◽

Vol 118 (21) ◽

pp. 3216-3216

Author(s):

M. Judith Peterschmitt ◽

Ravi S Kamath ◽

Elena Lukina ◽

Nora Watman ◽

Marta Dragosky ◽

...

Keyword(s):

Board Of Directors ◽

Research Funding ◽

Disease Type ◽

Phase 2 ◽

Z Score ◽

Advisory Committees ◽

Lytic Lesions ◽

Skeletal Complications ◽

Gaucher Disease Type 1

Abstract Abstract 3216 Introduction: In Gaucher disease type 1 (GD1), deficient lysosomal acid β-glucosidase leads to accumulation of undegraded glucosylceramide in lysosomes of tissue macrophages known as Gaucher cells. Skeletal complications are a major cause of morbidity and include bone marrow infiltration by Gaucher cells, osteopenia/osteoporosis, lytic lesions, fractures, avascular necrosis, and bone pain. Eliglustat, a novel, oral inhibitor of glucosylceramide synthase, is under investigation for the treatment of GD1. Objective: To report skeletal changes after 3 years of eliglustat therapy. Methods: This ongoing, open-label, uncontrolled, multicenter, Phase 2 clinical trial enrolled 26 adults with GD1 not on treatment for the previous 12 months, who had splenomegaly with thrombocytopenia and/or anemia. Study entry criteria also required that patients had no new pathologic bone involvement or bone crises within the preceding 12 months and not have used bisphosphonates during the previous 3 months. Changes from baseline were reported for centrally reviewed skeletal x-rays, dual-energy x-ray absorptiometry (DXA) and MRI assessments. Results: Of 26 enrolled patients, 19 completed 3 years of treatment. In 15 patients with evaluable DXA results at baseline and at 1, 2, and 3 years, mean lumbar spine BMD increased by 0.6±0.69 Z-score (baseline, −1.28), with greatest increases seen in osteoporotic patients. Mean femur BMD (T- and Z-score) remained normal through 3 years. Femur dark marrow on MRI, which reflects bone marrow infiltration by Gaucher cells, was reduced in 56% (10/18) or stable in 44% (8/18) of patients with findings at baseline. No bone crises or reductions in mobility occurred. On baseline radiographs, no patients had fractures, 42% (8/19 patients) had femoral lytic lesions, and 37% (7/19 patients) had bone infarcts. After 3 years, the lumbar spine and femurs showed no new lytic lesions, bone infarcts, fractures, or areas of osteonecrosis and no worsening of pre-existing lytic lesions or bone infarcts. One patient had worsening of asymptomatic osteonecrosis after 1 year noted retrospectively at baseline. Eliglustat was well-tolerated. Most adverse events (AEs) were mild and unrelated to treatment; the most common were viral infections (6 patients); urinary and upper respiratory tract infections (4 patients each); and headache, increased blood pressure, abdominal pain, diarrhea (3 patients each). Eight drug-related AEs, all mild, occurred in 6 patients. Conclusions: During 3 years of eliglustat treatment, radiologic monitoring showed improvement or stabilization of GD1 bony manifestations with no noted safety-related trends, suggesting that eliglustat may be a promising treatment for skeletal complications of GD1. Ongoing Phase 3 studies will provide more information on the bone effects of eliglustat. Disclosures: Peterschmitt: Genzyme: Employment. Kamath:Genzyme: Consultancy. Lukina:Genzyme: Honoraria. Watman:Genzyme: Membership on an entity's Board of Directors or advisory committees. Pastores:Genzyme: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Actelion: Research Funding; Amicus: Research Funding; Biomarin: Research Funding; Shire HGT: Research Funding; Protalix: Research Funding. Arreguin:Genzyme: Research Funding. Aguzzi:Genzyme: Employment. Ross:Genzyme: Employment. Puga:Genzyme: Employment. Rosenthal:Genzyme: Consultancy.

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Chemotherapy-Free Combination of Obinutuzumab and Ibrutinib in First LINE Treatment of Follicular Lymphoma. the Alternative Study By the German Low Grade Lymphoma Study Group (GLSG)

Blood ◽

10.1182/blood-2018-99-111692 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 448-448 ◽

Cited By ~ 2

Author(s):

Christian Schmidt ◽

Anna-Katharina Zoellner ◽

Vindi Jurinovic ◽

Martin Sökler ◽

Roswitha Forstpointner ◽

...

Keyword(s):

Board Of Directors ◽

Research Funding ◽

Tumor Burden ◽

Primary Efficacy ◽

Primary Efficacy Endpoint ◽

Advisory Committees ◽

Low Toxicity ◽

Grade 3 ◽

One Year ◽

Support Research

Abstract Background: The clinical course of follicular lymphoma (FL) is characterized by a slow progression over years with continuous relapses despite good response to initial treatment. The median overall survival is 10 to more than 15 years. Standard therapy for patients requiring treatment consists of an anti-CD 20 antibody combined with chemotherapy followed by antibody maintenance. With this combination a 1-year-PFS of 93% was seen in the GLSG-2000 trial (Hiddemann et al, Blood 2005). Because of the substantial side effects of chemotherapy such as infections, secondary malignancies and impairment of the stem cell reserve novel "chemotherapy-free" treatment approaches could substantially improve the treatment tolerability in FL. The BTK-inhibitor ibrutinib has demonstrated promising activity in patients with iNHL, CLL and MCL. Anticipating the recent reports on a superior activity of obinutuzumab as compared to rituximab in the GALLIUM trial (Marcus et al., NEJM 2017), the GLSG initiated a phase II study combining ibrutinib and obinutuzumab to explore the efficacy and safety of this "chemotherapy-free" alternative. Methods: ALTERNATIVE is a prospective multicenter single-arm phase 2 study of the combination of ibrutinib and obinutuzumab in 98 patients with previously untreated FL and a high tumor burden (defined by modified GELF criteria) in need of treatment. Induction comprises 6 cycles of obinutuzumab at a dose of 1000 mg by intravenous infusion on days 1, 8, 15 of cycle 1 and on day 1 of cycles 2-6 to be given every 21 days. Ibrutinib is administered orally at a dose of 560 mg once daily throughout all 6 cycles. In patients with at least partial response (defined by Cheson Response Criteria 2007) after the end of induction, maintenance with obinutuzumab (1000mg every 8 weeks) plus ibrutinib (560mg daily) is given for an additional 24 months. In patients remaining MRD positive at 30 months ibrutinib is continued for another 12 months in an extended maintenance setting without obinutuzumab. The primary efficacy endpoint is the rate of investigator-assessed PFS one year after registration. Response rates at end of induction, after one year and after end of maintenance, duration of response, percentage of progression during induction and maintenance, time to treatment failure, overall survival, duration of molecular remission in MRD negative patients and safety are key secondary endpoints. Results: 98 patients with advanced stage FL were included, The median age was 59 years (29-81), 60% were male and 40% had a high risk FLIPI, 90% stage III/IV disease and 10% were stage II with a high tumor burden. Response to in induction was 90% (87/97) with 85% (82/97) PR and 5% (5/97) CR. 5 patients (5%) progressed during induction. Of the 82 patients with PR after end of induction, 8 patients achieved a CR during the first 6 months of maintenance treatment. 95 patients were evaluable for the primary endpoint of 1-year-PFS and 76 patients (80%) remained alive and free of progression at this timepoint. 18 patients progressed in the first year, two of whom died due to progressive disease. One additional death was caused by a non-lymphoma related event. An MRD-marker was found in 65 patients. MRD at the end of induction was evaluable for 63 patients. 44 patients (70%) were MRD negative after induction treatment. Of the 42 patients with follow-up MRD peripheral blood or bone marrow samples, 35 (83%) were MRD negative one year after registration. Therapy was generally well tolerated. Most common adverse events were diarrhea in 30% of patients, rash in 25% and fatigue and nasopharyngitis (common cold) in 23% and 20%, respectively. Concerning hematotoxicity grade 3-4 neutropenia and thrombopenia were seen in 8% and 4% of patients, respectively. Severe (>=grade 3) infectious complications were rare (6% pneumonia/bronchitis, 2% sepsis, 7% other infections). Conclusions: The chemotherapy - free combination of ibrutinib and obinutuzumab showed high anti-lymyphoma activity with high overall response rates and a high proportion of MRD negativity at one year. While the combination of ibrutinib and obinutuzumab was associated with a low toxicity profile, the combination was inferior to the published results of conventional immunochemotherapies in terms of the primary efficacy endpoint (1-year-PFS). Further evaluations might demonstrate whether subgroups exist which particularly benefit clinically from this low toxicity regime. Figure Figure. Disclosures Schmidt: Celgene: Honoraria; Gilead: Honoraria, Other: Travel Grants; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel Grants. Buske:Bayer: Research Funding; Roche: Honoraria, Research Funding; Janssen: Honoraria, Research Funding. Viardot:Amgen: Consultancy; Gilead Kite: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; BMS: Consultancy, Honoraria; Roche: Consultancy, Honoraria. Keller:BMS: Consultancy; Roche: Consultancy; Takeda: Consultancy, Research Funding; Janssen-Cilag: Consultancy, Equity Ownership; MSD: Consultancy; Celgene: Research Funding. Graeven:Roche: Membership on an entity's Board of Directors or advisory committees; AbbVie: Honoraria. Marks:Merck: Honoraria; BMS: Honoraria; Servier: Honoraria. Hänel:Novartis: Honoraria; Roche: Honoraria; Amgen: Honoraria; Takeda: Honoraria. Liersch:Roche: Membership on an entity's Board of Directors or advisory committees; Abbvie: Honoraria. Dürig:Celgene: Honoraria; Roche: Honoraria, Speakers Bureau; Janssen: Consultancy, Honoraria. Hoster:Roche Pharma AG: Other: Travel support, Research Funding; F. Hoffman-La Roche: Other: Travel support, Research Funding. Unterhalt:F. Hoffman-La Roche: Other: Travel support. Hiddemann:Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; F. Hoffman-La Roche: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Bayer: Consultancy, Research Funding.

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Induction Therapy with Everolimus in Combination with DHAP (Dexamethasone, High-Dose AraC, Cisplatinum) in Patients with Relapsed or Refractory Classical Hodgkin Lymphoma: A Randomized, Placebo-Controlled Phase I/II Trial (HD-R3i)

Blood ◽

10.1182/blood-2018-99-117291 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 2912-2912 ◽

Cited By ~ 1

Author(s):

Bastian von Tresckow ◽

Andreas Hüttmann ◽

Vladan Vucinic ◽

Horst Mueller ◽

Annette Plütschow ◽

...

Keyword(s):

Placebo Group ◽

Phase I ◽

Board Of Directors ◽

Phase Ii ◽

Induction Therapy ◽

Research Funding ◽

High Dose ◽

Classical Hodgkin Lymphoma ◽

Advisory Committees ◽

Support Research

Abstract Introduction: Induction chemotherapy followed by BEAM high dose chemotherapy (HDCT) and autologous peripheral blood stem cell transplant (PBSC transplant) is standard of care for transplant-eligible patients with relapsed or refractory classical Hodgkin lymphoma (rrHL). However, approx. 50% of patients relapse and therefore, this strategy must be improved. As response to induction therapy is predictive of the outcome after HDCT, this trial aimed at improving the response to induction therapy by adding oral everolimus to time-intensified standard DHAP (Ever-DHAP). Methods: We included patients with histologically confirmed rrHL aged 18-60 years in this phase I/II trial. Dosage of everolimus was pre-determined in the phase I part with 10 mg/day given parallel to DHAP for 14 days within each of two cycles. The phase II part started as a randomized controlled trial comparing 50 patients in the everolimus group to 50 patients in a placebo group. The primary endpoint of the phase II part was the CT-based complete remission (CR-) rate after two cycles of Ever-DHAP. This CR-rate would be expected to be ≥ 40% if adding everolimus was effective. Secondary efficacy endpoints of the trial were PET-based CR-rate after two cycles of induction, progression-free and overall survival. Secondary feasibility endpoints were time to recovery, CTC-based adverse events, duration of induction therapy, discontinuation rates and the rates of successful PBSC collection. The trial was registered at ClinicalTrials.gov with ID NCT01453504. Results: From 7/2014 to 3/2018 we recruited a total of 59 patients in the phase II part. Because of poor recruitment the placebo group was closed in 9/2015 after 9 patients were randomized. These patients are analyzed in a descriptive way only. Of 50 patients in the everolimus group two were not evaluable because of retracting consent and not starting therapy; three additional patients discontinued Ever-DHAP because of toxicity. PBSC collection was successful in 37/39 documented patients receiving Ever-DHAP (95%). After two cycles of therapy we observed a CT-based CR in 12/45 patients of the everolimus group (27%) and in 2/9 patients of the placebo group (22%). A PET-based CR was achieved by 19/38 patients of the everolimus group (50%) and by 4/5 patients of the placebo group. In the everolimus group two patients had refractory disease (4%) and two died (4%), 3 and 4 months after starting but not related to Ever-DHAP. Final results and additional analyses will be presented. Conclusions: Adding everolimus to time-intensified DHAP is feasible, however, the Ever-DHAP regimen failed to show an improved efficacy. Disclosures von Tresckow: Novartis: Honoraria, Other: Travel support, Research Funding; Takeda: Honoraria, Other: Travel support, Research Funding; MSD: Honoraria, Other: Travel support, Research Funding. Hüttmann:Celgene: Other: Travel expenses; Roche: Other: Travel expenses. Viardot:Amgen: Consultancy; Roche: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; BMS: Consultancy, Honoraria; Gilead Kite: Consultancy, Honoraria. Topp:Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel, Research Funding; F. Hoffmann-La Roche Ltd: Membership on an entity's Board of Directors or advisory committees, Research Funding; Boehringer Ingelheim: Research Funding; Regeneron Pharmaceuticals, Inc.: Honoraria, Research Funding. Borchmann:Novartis: Consultancy, Honoraria.

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A Phase I/II Study of NMS-03592088, a FLT3, KIT and CSF1R Inhibitor, in Patients with Relapsed or Refractory AML or CMML

Blood ◽

10.1182/blood-2019-121954 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 3855-3855

Author(s):

Marina Ciomei ◽

Lucia Zanetta ◽

Federico Lussana ◽

Erika Ravelli ◽

Francesco Fiorentini ◽

...

Keyword(s):

Phase I ◽

Board Of Directors ◽

Phase Ii ◽

Research Funding ◽

Single Agent ◽

Dose Titration ◽

Advisory Committees ◽

Secondary Resistance ◽

Support Research ◽

Refractory Aml

Background: NMS-03592088 is a novel, potent inhibitor of the FLT3, CSF1R and KIT receptor tyrosine kinases (KD < 1 nM for all three targets). The compound demonstrated high preclinical efficacy following oral administration in all tested target-dependent tumor models, including those harboring kinase domain secondary resistance mutations, such us the FLT3 residue 691 gatekeeper mutation and the KIT residue 670 and exon 17 mutations. In a FLT3-ITD model of disseminated AML, efficacy observed following single agent treatment with NMS-03592088 was further significantly increased when administered in combination with cytarabine, with excellent tolerability. In preclinical studies conducted in non-human primates, a dose-related increase of circulating CSF1 levels was observed in association with the administration of NMS-03592088, consistent with in vivo inhibition of CSF1R by the compound, thus providing the opportunity for the use of CSF1 levels as a potential pharmacodynamic biomarker of CSF1R modulation in the clinical setting. All three targets of NMS-03592088 are relevant in different settings of hematologic malignancies and solid tumors. In particular, FLT3 mutations occur in approximately 30% of acute myeloid leukemia patients (AML), and are associated with a poor prognosis; KIT mutations are reported in patients with the core-binding factor (CBF) subtype of AML and the CSF1 and/or CSF1R genes are frequently expressed in AML blasts. Recent experimental evidence suggests a potential therapeutic rationale for CSF1R blockade in AML, possibly due to interference with microenvironmental support [Edwards DK et al, Blood, 2019, 133: 588]. Furthermore, chronic myelomonocytic leukemia (CMML) blasts express high levels of CSF1R and NMS-03592088 was able to effectively inhibit their proliferation, concomitant with the suppression of intracellular CSF1R dependent signalling. A clinical trial exploring safety, tolerability and efficacy of NMS-03592088 in patients with AML and CMML is therefore warranted. Trial design: This first-in-human study (EudraCT Number: 2018-002793-47) is designed as an open-label multicenter Phase I/II trial including patients with relapsed or refractory AML or CMML who have exhausted standard treatment options, or for whom standard therapy is considered unsuitable. The study is designed to characterize the safety, tolerability, pharmacokinetics, pharmacodynamics and to explore the preliminary anticancer activity of NMS-03592088 administered orally as single agent once daily for 21 consecutive days, followed by a 7-day break within a 28 day cycle. The study includes an initial conventional phase I part with an accelerated dose titration design in subsequent cohorts of 3+3 patients aimed at defining the maximal tolerated dose (MTD) and the recommended phase 2 dose (RP2D), followed by a limited dose expansion to confirm the RP2D. Once the RP2D is confirmed, a single-stage exploratory Phase II part will start comprising two parallel cohorts, one cohort will consist of AML FLT3 mutated patients and one of patients with CMML. Patients previously treated with FLT3 inhibitors are allowed to participate. The primary endpoint of the Phase II portion of the study is Overall Response Rate. Efficacy will be assessed according to standard criteria [Döhner H et al, Blood 2017, 129: 424; Savona MR et al., Blood, 2015, 125: 1857]. Exploratory endpoints are included to evaluate the potential effects of treatment with NMS-03592088 on circulating levels of CSF1 in plasma, the potential correlation of cellular CSF1R expression levels with clinical outcome in both AML and CMML, and the mutational status of a panel of leukemia-related genes, not limited to FLT3. The Phase I part started in Italy in March, 2019 and is currently ongoing. Disclosures Ciomei: NMS: Employment. Zanetta:Clioss: Employment. Fiorentini:Accelera: Employment. Bosotti:NMS: Employment. Ardini:NMS: Employment. Lombardi Borgia:NMS: Employment. Pulci:Accelera: Employment. Gatto:Clioss: Employment. Di Sanzo:Clioss: Employment. Colajori:Clioss: Consultancy. Davite:Clioss: Employment. Galvani:NMS: Employment. Gan:NMS: Employment. Rossi:Janssen: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Amgen: Membership on an entity's Board of Directors or advisory committees; Gilead: Membership on an entity's Board of Directors or advisory committees; Sanofi: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees; Pfizer: Membership on an entity's Board of Directors or advisory committees; Jazz: Membership on an entity's Board of Directors or advisory committees; Astellas: Membership on an entity's Board of Directors or advisory committees; Novartis: Honoraria; Mundipharma: Honoraria; BMS: Honoraria; Sandoz: Honoraria; Daiichi-Sankyo: Consultancy; Roche: Membership on an entity's Board of Directors or advisory committees. Rambaldi:Jazz: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau, travel support; Novartis: Membership on an entity's Board of Directors or advisory committees, Other: travel support, Speakers Bureau; Celgene: Membership on an entity's Board of Directors or advisory committees, Other: travel support, Speakers Bureau; Roche: Membership on an entity's Board of Directors or advisory committees, Other: travel support, Research Funding, Speakers Bureau; Amgen: Membership on an entity's Board of Directors or advisory committees, Other: travel support, Research Funding, Speakers Bureau; Pfizer: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Italfarmaco: Membership on an entity's Board of Directors or advisory committees, Other: travel support, Research Funding, Speakers Bureau; Gilead: Membership on an entity's Board of Directors or advisory committees, Other: travel support, Speakers Bureau; Omeros: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Isacchi:NMS: Employment.

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Clinical Study to Investigate the Efficacy and Safety of Wilate during Prophylaxis in Previously Treated Patients with Von Willebrand Disease (VWD)

Blood ◽

10.1182/blood-2019-128339 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 4931-4931

Author(s):

Robert F. Sidonio ◽

Bruce A. Schwartz

Keyword(s):

Board Of Directors ◽

Research Funding ◽

Prophylactic Treatment ◽

Von Willebrand Disease ◽

Advisory Committees ◽

Previously Treated Patients ◽

Previously Treated ◽

Type 3 ◽

Von Willebrand

Background: Inherited von Willebrand disease (VWD) is the most common inherited hemorrhagic disorder, with an estimated prevalence of 1 in every 100 individuals. Type 1 and type 3 (the most severe form) are characterized by a quantitative deficiency of von Willebrand factor (VWF) and type 2 arises from a qualitative deficiency of VWF. Treatment of VWD depends on the type and severity of the disease. Severe bleeding is reported in patients with all subtypes, leading to progressive joint disease as well as diminished quality of life (QoL). VWF/factor VIII (FVIII) concentrates have become the mainstay of VWD treatment for these patients with severe disease or for those patients in whom other treatments (e.g., desmopressin) are ineffective or contraindicated but this is broadly applicable only for on demand treatment. Aims: The primary objective of this study is to determine the efficacy of VWF/FVIIII concentrate in the prophylactic treatment of previously treated patients with type 3, type 2 (except 2N), or severe type 1 VWD. Secondary objectives of this study will be to collect data to 1) Assess the VWF:Ac and VWF:Ag incremental IVR of VWF/FVIIII concentrate over time and, 2) Assess the safety and tolerability of VWF/FVIIII concentrate in this indication. The study will also examine, the efficacy of VWF/FVIIII concentratein the treatment of breakthrough bleeding episodes (BEs), and in surgical prophylaxis, as well as the QoL during prophylaxis with VWF/FVIIII concentrate. Methods: The study is planned to enroll 28 patients aged ≥6 years and with VWD type 1, 2A, 2B, 2M, or 3. Eligible patients must be receiving on-demand treatment with a VWF-containing product, with at least 1, and an average of ≥2, documented spontaneous BEs per month in the preceding 6 months requiring treatment with a VWF-containing product. This will be assessed as part of a run-in observational study to collect the bleeding profile prior to the start of prophylaxis. From the beginning of the study, patients will receive prophylactic treatment with VWF/FVIIII concentrate for 12 months and record all BEs in a patient diary. Based on these data, the frequency of BEs and the annualized bleeding rate (ABR) under prophylactic treatment will be calculated. Treatment efficacy of BEs will be assessed by the patient (together with the investigator in case of on-site treatment) using a 4-point scale (excellent, good, moderate, none) In patients that undergo surgeries, efficacy of VWF/FVIIII concentratewill be assessed at the end of surgery by the surgeon and at the end of the postoperative period by the haematologist. In both cases, predefined assessment criteria will be used. In addition, an overall assessment of efficacy will be made at the end of the postoperative period by the investigator. Results: Data will be monitored on an ongoing basis and the study is expected to end Q2 2021. Conclusions: Prophylactic treatment in other congenital bleeding disorders is widely accepted as the standard of care to prevent bleeding and preserve QoL in patients but to date, this form of treatment in VWD is not well characterized. This study will provide data on the efficacy of prophylactic treatment in reducing the rate of bleeding and on the impact of prophylaxis on the QoL in VWD patients. Disclosures Sidonio: Genetech: Membership on an entity's Board of Directors or advisory committees, Research Funding; Takeda-Shire: Membership on an entity's Board of Directors or advisory committees, Research Funding; Biomarin: Membership on an entity's Board of Directors or advisory committees; Octapharma: Membership on an entity's Board of Directors or advisory committees, Research Funding; Bioverativ: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novo Nordisk: Membership on an entity's Board of Directors or advisory committees; Grifols: Membership on an entity's Board of Directors or advisory committees, Research Funding; Kedrion: Research Funding; Uniqure: Membership on an entity's Board of Directors or advisory committees. Schwartz:Octapharma: Employment.

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The Impact of gain1q on Mutational Structure and Clonal Evolution in a Uniformly Treated High-Risk Series of Patients at First Relapse

Blood ◽

10.1182/blood-2021-152123 ◽

2021 ◽

Vol 138 (Supplement 1) ◽

pp. 2683-2683

Author(s):

John R Jones ◽

Charlotte Pawlyn ◽

Niels Weinhold ◽

Timothy Cody Ashby ◽

Brian A Walker ◽

...

Keyword(s):

High Risk ◽

Board Of Directors ◽

Research Funding ◽

Clonal Evolution ◽

Suppressor Gene ◽

Tumour Suppressor Gene ◽

Mutational Load ◽

Advisory Committees ◽

The Impact ◽

Support Research

Abstract Introduction In Multiple Myeloma (MM) the emergence of treatment resistant clones is a characteristic feature of relapse and this is particularly so for high-risk cases. A key driver event mediating progression, risk status and relapse is gain(1q) (1q+). We report on the impact of 1q+ on the genetic profile seen at first relapse in a uniformly treated, newly diagnosed series of 56 patients enrolled to the NCRI Myeloma XI Trial. Methods We included 56 high risk patients, defined as relapse within 30 months of maintenance randomisation (median 19 months, range 8-51). Of the 56 patients, 30 received lenalidomide maintenance and 26 were observed. Whole exome sequencing was conducted at presentation and relapse to a median depth of 122x for tumour samples and 58x for controls. Libraries were prepared using the SureSelectQXT sample prep kit and SureSelect Clinical Research Exome kit. MuTect was used to determine gene variants and SciClone clustering was undertaken to map mutation variant allele frequencies. MANTA was used to determine translocations and Sequenza for copy number aberrations. Clonal structure and mechanisms of clonal evolution were assessed using kernel density estimation of the cancer clonal fraction for all mutations. Wilcoxon matched-pairs signed rank tests (2-sided) were used to determine the significance between paired data sets, including mutational load. Fishers exact test was used to determine the difference between two nominal variables. Results We looked at mutational, structural and clonal evolution events in all patients based on 1q+ status at relapse. At diagnosis, 34% (19/56) patients had evidence of 1q+, increasing to 46% (26/56) at relapse, with all patients harbouring 1q+ at presentation having the lesion at relapse. There was a significantly higher non-synonymous mutational load at relapse in patients with 1q+, 107 vs 126 (p=0.047), compared to those without 1q+, 36 vs 44 (p=0.140). Twenty two genes known to be significant in MM and mutations within the genes known to be important in IMiD mechanism of action were reviewed. Of the patients with 1q+, 92% (24/26) had at least one mutation during the course of the disease, compared to 77% in those without 1q+ (p=0.15). The impact on tumour suppressor gene regions including deletions of chromosome 1p, 13, 14 and 17p was analyzed. Of the patients with 1q+, 77% (20/26) of patients had a deletion of one of these regions during the disease course, compared to 57% (17/30) of patients without 1q+ (p=0.16). At relapse a change in the profile of these lesions was noted in 23% (6/26) patients with 1q+, compared to 20% (6/30) patients without 1q+ (p=1). Translocations involving MYC (t MYC) were also determined and found in 27% (7/26) of patients with 1q+ and 27% (8/30) of patients without (p=1). As with 1q+, t MYC was always preserved at relapse. Mechanisms of evolution leading to relapse were established for all patients. Branching and linear evolution predominated, noted to be the mechanism leading to relapse in 88% (23/26) patients with 1q+ and 83% (25/30) without (p0.71). Stable evolution was noted in the remaining patients. 1q+ occurring as a new event at relapse was associated with branching or linear evolution in all patients (n=7), consistent with a change in clonal structure. Conclusion These data reveal that 1q+ is conserved throughout the disease course, suggesting it imparts a survival advantage and treatment resistant phenotype to the clone(s) containing it. The presence of 1q+ is associated with a significant increase in mutational load at relapse and a greater incidence of tumour suppressor gene structural deletions, mechanisms that may contribute to clonal evolution and therapeutic escape. Disclosures Jones: BMS/Celgene: Other: Conference fees; Janssen: Honoraria. Pawlyn: Celgene / BMS: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria; Sanofi: Honoraria, Membership on an entity's Board of Directors or advisory committees. Weinhold: Sanofi: Honoraria. Walker: Sanofi: Speakers Bureau; Bristol Myers Squibb: Research Funding. Cairns: Merck Sharpe and Dohme: Research Funding; Amgen: Research Funding; Takeda: Research Funding; Celgene / BMS: Other: travel support, Research Funding. Kaiser: AbbVie: Consultancy; Seattle Genetics: Consultancy; BMS/Celgene: Consultancy, Other: Travel support, Research Funding; Amgen: Honoraria; Karyopharm: Consultancy, Research Funding; Pfizer: Consultancy; Janssen: Consultancy, Other: Educational support, Research Funding; GSK: Consultancy; Takeda: Consultancy, Other: Educational support. Cook: Pfizer: Consultancy, Honoraria; Karyopharm: Consultancy, Honoraria; BMS: Consultancy, Honoraria, Research Funding; Sanofi: Consultancy, Honoraria; Oncopeptides: Consultancy, Honoraria; Janssen: Consultancy, Honoraria, Research Funding; Takeda: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Roche: Consultancy, Honoraria. Drayson: Abingdon Health: Current holder of individual stocks in a privately-held company. Jackson: oncopeptides: Consultancy; takeda: Consultancy, Honoraria, Research Funding, Speakers Bureau; GSK: Consultancy, Honoraria, Speakers Bureau; J and J: Consultancy, Honoraria, Speakers Bureau; celgene BMS: Consultancy, Honoraria, Research Funding, Speakers Bureau; amgen: Consultancy, Honoraria, Speakers Bureau; Sanofi: Honoraria, Speakers Bureau. Davies: BMS: Consultancy, Honoraria; Takeda: Consultancy, Honoraria; Abbvie: Consultancy, Honoraria; Janssen: Consultancy, Honoraria; Amgen: Consultancy, Honoraria; Roche: Consultancy, Honoraria. Morgan: BMS: Membership on an entity's Board of Directors or advisory committees; Jansen: Membership on an entity's Board of Directors or advisory committees; Karyopharm: Membership on an entity's Board of Directors or advisory committees; Oncopeptides: Membership on an entity's Board of Directors or advisory committees; GSK: Membership on an entity's Board of Directors or advisory committees.

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